Agents promoting laminin production in skin cells

ABSTRACT

Disclosed are compositions for promoting the production of laminin 5 in epidermic cells which contain a lysophospholipid having a C 14-22  fatty acid residue or a preparation derived from soybeans as an active ingredient and which can be used in the fields of cosmetics and dermatology. These compositions may be used especially for the purpose of potentiating the skin.

TECHNICAL FIELD

This invention relates to agents for promoting the production of laminin5 in epidermic cells and skin potentiating compositions useful in thefields of cosmetics and dermatology. More particularly, this inventionrelates to the use of specific lysophospholipids and soybean-derivedpreparations for the purpose of potentiating skin functions through themedium of the promotion of laminin 5 production in epidermic cells.

BACKGROUND ART

In the fields of cosmetics and dermatology, a wide variety of means havebeen proposed and tried to mitigate or cure the damage of the skin dueto the influence of an external environment (e.g., exposure to sunlight)and aging. For example, main skin changes caused by aging includewrinkle formation, hardening and reduced elasticity.

In connection with the causes of such changes, chief interest isdirected not to the epidermis of the skin, but rather to the functionsof collagen fibers and elastic fibers consisting of collagen andglycosaminoglycans and present in the dermis forming the substratum ofthe epidermis. As means for preventing or correcting such changes, theuse of a hydroxycarboxylic acid (e.g., Japanese Patent No. 2,533,339)and the use of a lysophospholipid [Japanese Patent Laid-Open No.67621/'96 or Journal of Japan Oil Chemists' Society, Vol. 46, No. 9(1997), pp. 13-19] have been proposed. In the former patent, it issuggested that the stratum corneum and wrinkles can be exterminated bypreventing a loss of collagen fibers. On the other hand, it is suggestedin the latter patent that lysophospholipids enhance the ability of humanfibroblasts to produce a glycosaminoglycan (specifically, hyaluronicacid) and hence exhibit a skin beautifying effect. It is also suggestedthat lysophospholipids exert little influence on the synthesis ofcollagenous protein in skin fibroblasts (see the aforementioned Journalof Japan Oil Chemists' Society).

In the year 1991, laminin 5 was discovered by Dr. Burgeson (Rousselle etal., J. Cell Biol., 114, 567, 1991). Subsequently, the presence of anautoimmune disease to laminin 5 (Fine et al., J. Am. Acad. Dermatol.,24, 119, 1991) and a serious disease caused by the genetic deficiency oflaminin 5 (Aberdam et al., Nat. Genet., 6, 299, 1994) was discovered. Inthis disease, the formation of blisters was observed at thedermal-epidermal junction, revealing that laminin 5 is an indispensablecomponent for the adhesion of them. It is known that compounds of thelaminin family are synthesized in various types of cells. Among them,laminin 5 is a principal component of the structure (called the basementmembrane) located at the dermal-epidermal junction, and composed ofvarious glycoproteins and proteoglycans. Moreover, laminin 5 has anactivity for promoting the adhesion of epidermic cells and therebyfunctions to bind epidermic cells directly to the basement membrane(Rousselle et al., J. Cell Biol., 114, 567, 1991). Furthermore, it isalso known that the adhesion of epithelial cells to the basementmembrane components is important in maintaining their cellular function(Schmidhauser et al., Proc. Natn. Acad. U.S.A., 87, 9, 118, 1990).Furthermore, in the skin of old persons, duplication of the basementmembrane (Lavker et al., J. Invest. Dermatol., 73, 59, 1979) and athickening of the basement membrane and a decrease of IV type collageconstituting it (Vazquez, Maturitas, 25, 209, 1996) have been reported.Thus, there seems to be a possibility that structural changes of thebasement membrane may be responsible for hypofunction of the skin in oldpersons.

As to Japanese Patent Laid-Open No. 67621/'96 and Journal of Japan OilChemists' Society, Vol. 46, No. 9 (1997), pp. 13-19, the latter morespecifically explains the contents of the former with the aid ofadditional experimental data. According to the latter (i.e., theaforementioned Journal of Japan Oil Chemists' Society), it is suggestedthat, among the lysophospholipids represented by the formula

the compound in which R is a C₁₂ saturated fatty acid residue promotesthe above-described production of hyaluronic acid, but the compounds inwhich R is a C₁₄ or higher saturated fatty acid residue inhibits itsproduction strongly.

DISCLOSURE OF THE INVENTION

On the basis of the above-described findings, the present inventors havesearched for substances capable of improving the function of the skin onthe assumption that the firm binding of epidermic basal cells to thebasement membrane is essential for the manifestation of normal skinfunctions and that it is more important to promote the production oflaminin 5 in the corium and the like, rather than the production ofcollagen and hyaluronic acid.

As a result, the present inventors have now found that, in contrast tothe aforementioned Journal of Japan Oil Chemists' Society suggestingthat the compounds of the above formula in which R is a C₁₄ or highersaturated fatty acid residue strongly inhibits the production ofhyaluronic acid in human skin fibroblasts, the compounds of the aboveformula in which R is a C₁₄₋₂₂ fatty acid residue optionally having upto six unsaturated double bonds, and soybean-derived preparations thatare likely to contain such compounds or their precursors, are moreeffective in promoting the above-described production of laminin 5 andthereby potentiating the skin.

Accordingly, the present invention relates to a composition forpromoting the production of laminin 5 in epidermic cells which containsa compound of the following formula (I) or a mixture of two or more suchcompounds, as an active ingredient.

wherein R is a C₁₄₋₂₂ fatty acid residue optionally having up to sixunsaturated double bonds, A is a hydrogen atom or a —CH₂CH₂N⁺(CH₃)₃group, and M is a hydrogen atom or an alkali metal atom, but is ahydrogen atom when A is a —CH₂CH₂N⁺(CH₃)₃ group.

In other embodiments, the present invention relates to a method forpromoting the production of laminin 5 in epidermic cells byadministering the composition to the skin of a mammal, and the use of acompound of the following formula (I) as an active ingredient of acomposition for promoting the production of laminin 5.

In further embodiments, the present invention also relates to acomposition for promoting the production of laminin 5 which contains asoybean-derived preparation that is likely to contain a compound of theabove formula (I) or its precursor, as an active ingredient, a methodfor promoting the production of laminin 5 in epidermic cells byadministering the composition to the skin of a mammal, and the use ofthe soybean-derived preparation as an active ingredient of a compositionfor promoting the production of laminin 5.

DETAILED DESCRIPTION OF THE INVENTION

According to the present invention, the compounds of the above formula(I) wherein R is a C₁₄₋₂₂ fatty acid residue, which may strongly inhibitthe production of hyaluronic acid in human skin fibroblasts as suggestedin the aforementioned Journal of Japan Oil Chemists' Society, andsoybean-derived preparations that are likely to contain such compounds,can be effectively used especially for the purpose of promoting theproduction of laminin 5 and/or potentiating the skin through the mediumof such action.

The mammals in which it is contemplated to promote the production oflaminin 5 and/or potentiate the skin through the medium of such actionaccording to the present invention include, but are not limited to,human beings, pet animals (e.g., dogs, cats, rats and guinea pigs) anddomestic animals (e.g., cattle, pigs and sheep).

The compounds of the above formula (I), which are used in the presentinvention, are compounds known generically as lysophospholipids. Thesecompounds may be directly or indirectly obtained from a wide variety ofbiological sources, but may also be chemically synthesized.

R in the above formula (I) is a radical selected from among C₁₄₋₂₂ fattyacid residues (acyl groups). Specifically, these acyl groups includeresidues derived from saturated fatty acids such as myristic acid (C₁₄),pentadecanoic acid (C₅), palmitic acid (C₆), margaric acid (C₁₇),stearic acid (C₈), nonadecanoic acid (C₁₉), arachidic acid (C₂₀),heneicosanoic acid (C₂₁) and behenic acid (C₂₂). Of these, the preferredresidues are those derived from C₁₄, C₁₆, C₁₈, C₂₀ and C₂₂ saturatedfatty acids which are abundantly found especially in biological sources.Moreover, from the viewpoint of activity for promoting the production oflaminin 5, the residues derived from C₆ and C₁₈ saturated fatty acidsare preferred.

Furthermore, R can be an acyl group comprising any of theabove-described fatty acid residues in which up to six unsaturateddouble bonds are present. The positions and number of double bonds insuch fatty acid residues should preferably correspond to those in thefatty acid residues of phospholipids or lysophospholipids obtainablefrom biological sources. However, no limitation is placed thereon, solong as the number of double bonds is six or less and is not contrary tothe purpose of the present invention. Among the fatty acids forming suchacyl groups, oleic acid (C₁₈: (Z)-9-octadecenoic acid), elaidic acid(C₁₈: (E)-9-octadecenoic acid), linolic acid (C₁₈: 9,12-octadecadienoicacid), α-linolenic acid (C₁₈: 9,12,15-octadecatrienoic acid),β-linolenic acid (C₁₈: 6,9,12-octadecatrienoic acid),5,8,11-icosatrienoic acid (C₂₀), 5,8,11,14-eicosatetraenoic acid (C₂₀),4,8,12,15,19-docosapentaenoic acid (C₂₂) and4,7,10,13,16,19-docosahexaenoic acid (C₂₂) are especially preferred.

A in formula (I) can be a hydrogen atom or a —CH₂CH₂N⁺(CH₃)₃ group. WhenA is the former, the compound is the so-called lysophosphatidic acid,and when A is the latter, the compound is the so-calledlysophosphatidylcholine. In the case of lysophosphatidic acid, it may bein the form of a salt in which M in formula (I) is an alkali metal.Preferred examples of this alkali metal include potassium, sodium andlithium.

In the present invention, the above-described compounds of formula (I)may be used alone or in a mixture of two or more. Such mixtures may beformed by combining two or more compounds in which R is any of theabove-described preferred groups. Alternatively, alysophosphatidylcholine mixture may be prepared by obtaining aphospholipid from a biological source [in particular, seeds of thesoybean (Glycine max Merril)], hydrolyzing it with phospholipase A₂known per se to form a lysophospholipid, hydrolyzing it withphospholipase D or by chemical means as required, and purifying it, andmay be used as such. Such lysophosphatidylcholine mixtures include, forexample, Reagent Lysolecithin Prepared from Soybeans (Wako Pure ChemicalIndustries Ltd.; Catalog No. 120-00832), Lysolecithin Kyowa (Iwase CosfaCo., Ltd.), Sunlecithin WL-25 (Taiyo Chemical Industry Co., Ltd.),Blendmax (Central Soya) and LIPIDURE NC-22 (Nippon Oil & Fats Co.,Ltd.), which may be used as such or, if necessary, after furtherpurification.

Alternatively, a soybean-derived preparation which has not necessarilybeen hydrolyzed or purified as described above, but can promote theproduction of laminin 5, may be used as the active ingredient of thepresent invention. Such soybean-derived preparations include groundsoybeans as such, and extracts obtained by extracting ground soybeanswith a suitable solvent. Such extracts include, but are not limited to,ones obtained by extraction with a water-containing lower alcohol, loweralcohol, lower alkyl ketone, lower alkyl ether, polyhydric alcohol or amixture of two or more such solvents.

Specific examples of useful water-containing lower alcohols includemethanol, ethanol, isopropanol and n-propanol containing less than 50%by weight, preferably 5 to 40% by weight, of water; specific examples ofuseful lower alcohols include methanol, ethanol, isopropanol,n-propanol, n-butanol and, sec-butanol, containing essentially no water;and specific examples of useful lower alkyl ketones include acetone,methyl ethyl ketone, butyl methyl ketone and isobutyl methyl ketone.

Specific examples of useful di-lower alkyl ethers include diethyl ether,di-isopropyl ether, di-n-propyl ether, di-n-butyl ether, n-butyl methylether and n-butyl ethyl ether; and specific examples of useful loweralkyl esters of carboxylic acids include ethyl acetate and butylacetate.

Specific examples of useful polyhydric alcohols include 1,3-butanediol,propylene glycol, glycerol and diglycerol.

Moreover, there may also be used preparations obtained by extractingsoybeans with an extraction solvent selected from hydrocarbons such ascyclohexane, toluene and xylene; and halogenated hydrocarbons such asdichloromethane and dichloroethane, so long as they meet the purpose ofthe present invention.

Soybeans or ground soybeans are extracted with a solvent as describedabove according to any of various per se known methods, and theresulting extract is used, either as such or after drying it andredissolving it in a suitable solvent, to measure its activity for theproduction of laminin 5 as will be described later. Thus, thepreparation used in the present invention can be selected. Theextraction may be carried out according to any of various procedurescommonly employed in solid-liquid extraction, for example, by soakingoptionally dried soybeans or ground soybeans in any of theabove-described solvents and allowing it to stand or shaking it on ashaker. Although the extraction temperature may be suitably chosenaccording to the solvent system used, it usually ranges from 5° C. tothe boiling point of the solvent used and preferably up to 60° C. As tothe extraction time, the optimum condition for the preparation used inthe present invention may vary according to the solvent used and thetemperature. However, it would be easy for those skilled in the art todetermine the extraction time by measuring the activity for theproduction of laminin 5 as will be described layer.

The preferred extraction procedure comprises using 70 to 95% methanol orethanol (having a water content of 30 to 5% and, if necessary, adjustedto pH 3-8), absolute methanol, absolute ethanol, acetone and diethylether or 1,3-butanediol, or propylene glycol, and soaking soybeans orground soybeans therein at room temperature (i.e., 15 to 30° C.) toextract desired components. The extract thus obtained may be used in thepresent invention, either as such or after being further diluted withethanol, or the dried extract may be used in the present invention,either as such or after being redissolved in ethanol.

Commercially available, typical examples of such soybean-derivedpreparations include soybean saponin [Saponin, Prepared from Soybeans;Wako Pure Chemical Industries Ltd., Tokyo (Catalog No. 190-08852)] andsoybean lecithin [Lecithin, Prepared from Soybeans; Wako Pure ChemicalIndustries Ltd., Tokyo (Catalog No. 120-00832)].

The target cells for the present invention are different from the targetcells for the above-described production of hyaluronic acid. Moreover,as specified by formula (I), the compounds intended for use in thepresent invention have not necessarily the same usable range oreffective concentration as the compounds used for promoting theproduction of hyaluronic acid.

According to the present invention, there is provided a skinpotentiating composition which contains, as an active ingredient, acompound of the above formula (I) or a mixture of two or more suchcompounds, or a preparation derived from soybeans, at a concentrationsufficient to promote the production of laminin 5 in epidermic cells.The expression “skin potentiating” means to improve hypofunction of theskin associate, for example, with structural changes of the basementmembrane due to aging or the like, more specifically, wrinkled skin andhardened skin.

The aforesaid “concentration sufficient to promote the production oflaminin 5” may vary according to the type of the compound of formula (I)or soybean-derived preparation used, other ingredients used to preparethe composition, the dosage form of the composition, and its applicationtime. Accordingly, the optimum concentration of a compound of formula(I) or a mixture of such compounds needs to be determined for each givencomposition. It would be easy for those skilled in the art to determinethe optimum concentration by referring to this specification (inparticular, the examples which will be given later) and carrying out asmall-scale experiment as required.

However, when a compound of formula (I) or a mixture of such compoundsis used, its concentration may generally be in the range of 0.0001 to60% by weight, preferably 0.01 to 60% by weight, based on the totalweight of the composition. On the other hand, when a soybean-derivedpreparation is used, its optimum amount used may generally varyaccording to the method of preparation from soybeans and, therefore,cannot be critically defined. However, when an extract obtained byextracting dry ground soybeans with water-containing methanol or ethanolor with a mixture of ethanol or acetone and diethyl ether is used as thepreparation, its content on a dry solid basis may be in the range of0.0001 to 20% by weight based on the total weight of the composition.

The composition of the present invention, which has been prepared in theabove-described manner, can promote the production of laminin 5 in theskin by administering it topically to a mammal (e.g., by applying it tothe skin). The topical administration may be carried out several times aday or at regular intervals (e.g., at intervals of several days)according to the intended purpose. The frequency of administration andthe amount of active ingredient administered at a time can be determinedwith consideration for the results of tests on experimental animals aswill be described later, or the results of the actual use in volunteers(because the compositions of the present invention exhibit essentiallyno toxicity to mammals).

Moreover, especially when the active ingredient comprises a compound offormula (I), the compositions of the present invention may beadministered orally or parenterally.

The skin potentiating compositions of the present invention may takevarious forms such as an aqueous solution, an oil solution, othersolutions, a milky lotion, cream, gel, a suspension, microcapsules,powder, granules, capsules and a solid. After they are made into theseforms according to the per se known methods, the resulting preparationsincluding lotions, milky lotions, creams, ointments, plasters,poultices, aerosols, injections, internal preparations (i.e., tablets,powders, granules, pills, syrups, troches, etc.), suppositories and thelike may be applied to, sticked on, sprayed on, injected into, drunkinto, or inserted into the body. Among these preparations, externalpreparations for the skin such as lotions, milky lotions, creams,ointments, plasters, poultices and aerosols are considered to be dosageforms suitable for the compositions of the present invention. Theabove-described “external preparations for the skin” include drugs,quasi drugs and cosmetics, and the same shall apply hereinafter.

The compositions of the present invention may suitably containexcipients and other additives (e.g., perfumes) which are commonly usedin the preparation of such compositions, as well as fats and oils,surfactants, antiseptics, sequestering agents, water-soluble polymers,thickeners, powder ingredients, ultraviolet protectants, humectants,active ingredients, antioxidants, pH regulators, detergents, desiccants,emulsifiers and the like. When these various ingredients areincorporated into the skin potentiating compositions of the presentinvention, they must be added to such an extent as not to detract fromthe desired effects of the present invention.

The aforesaid fats and oils include liquid oils, solid fats, waxes,hydrocarbon oils, higher fatty acids, higher alcohols, synthetic esteroils and silicones.

Specifically, useful liquid oils include avocado oil, camellia oil,evening primrose oil, turtle oil, macadamia nut oil, corn oil, mink oil,olive oil, rapeseed oil, egg yolk oil, sesame oil, persic oil, wheatgerm oil, sasanqua oil, castor oil, linseed oil, safflower oil,cottonseed oil, perilla oil, soybean oil, peanut oil, tea seed oil, kayaoil, rice bran oil, Chinese tung oil, Japanese tung oil, jojoba oil,germ oil, triglycerol, glycerol trioctanoate, glycerol triisopalmitateand the like; useful solid fats include cacao butter, coconut oil, horsefat, hardened coconut oil, palm oil, beef tallow, mutton tallow,hardened beef tallow, palm kernel oil, lard, beef bone fat, haze kerneloil, hardened oil, beef foot fat, neat's foot oil, Japan wax, hardenedcastor oil and the like; useful waxes include beeswax, candelilla wax,cotton wax, carnauba wax, bayberry wax, Chinese wax, spermaceti, montanwax, rice bran wax, lanolin, kapok wax, lanolin acetate, liquid lanolin,sugarcane wax, lanolin fatty acid isopropyl ester, hexyl laurate,reduced lanolin, jojoba wax, hard lanolin, shellac wax, POE lanolinalcohol ether, POE lanolin alcohol acetate, POE cholesterol ether,lanolin fatty acid polyethylene glycol ester, POE hydrogenated lanolinalcohol ether and the like; and useful hydrocarbon oils include liquidparaffin, ozokerite, squalene, pristane, paraffin, ceresine, squalene,vaseline, microcrystalline wax and the like.

Useful higher fatty acids include, for example, lauric acid, myristicacid, palmitic acid, stearic acid, behenic acid, oleic acid,12-hydroxystearic acid, undecylenic acid, tallic acid, isostearic acid,linolic acid, linoleic acid, eicosapentaenoic acid (EPA) anddocosahexaenoic acid (DHA).

Useful higher alcohols include, for example, straight-chain alcoholssuch as lauryl alcohol, cetyl alcohol, stearyl alcohol, behenyl alcohol,myristyl alcohol, oleyl alcohol and cetostearyl alcohol; andbranched-chain alcohols such as monostearyl glycerol ether (batylalcohol), 2-decyltetradecynol, lanolin alcohol, cholesterol,phytosterol, hexyldodecanol, isostearyl alcohol and octyldodecanol.

Useful synthetic ester oils include, for example, isopropyl myristate,cetyl octanoate, octyldodecyl myristate, isopropyl palmitate, butylstearate, hexyl laurate, myristyl myristate, decyl oleate, hexyldecyldimethyloctanoate, cetyl lactate, myristyl lactate, lanolin acetate,isocetyl stearate, isocetyl isostearate, cholesteryl 12-hydroxystearate,ethylene glycol di-2-ethylhexylate, dipentaerythritol fatty acid esters,N-alkyl glycol monoisostearate, neopentyl glycol dicaprate, diisostearylmalate, glycerol di-2-heptylundecanoate, trimethylolpropanetri-2-ethylhexylate, trimethylolpropane triisostearate, pentaerythritoltetra-2-ethylhexylate, glycerol tri-2-ethylhexylate, trimethylolpropanetriisostearate, cetyl 2-ethylhexanoate, 2-ethylhexyl palmitate, glyceroltrimyristate, tri-2-heptylundecanoic acid glyceride, castor oil fattyacid methyl ester, oleyl oleate, cetostearyl alcohol, acetoglyceride,2-heptylundecyl palmitate, diisobutyl adipate, 2-octyldodecylN-lauroyl-L-glutamate, di-2-heptylundecyl adipate, ethyl laurate,di-2-ethylhexyl sebacate, 2-hexyldecyl myristate, 2-hexyldecylpalmitate, 2-hexyldecyl adipate, diisopropyl sebacate, 2-hexyldecylsuccinate, ethyl acetate, butyl acetate, amyl acetate and triethylcitrate.

Useful silicones include chain polysiloxanes such as dimethylpolysiloxane, methyl phenyl polysiloxane and methyl hydrogenpolysiloxane; cyclic polysiloxanes such as decamethyl polysiloxane,dodecamethyl polysiloxane and tetramethyl tetrahydrogen polysiloxane;silicone resin having a three-dimensional network structure; siliconerubber; and the like.

Since the compounds of formula (I) which are used in the presentinvention have surface activity in themselves, it is usually unnecessaryto use a surfactant. However, depending on the type of the composition,other anionic surfactants, cationic surfactants, ampholytic surfactantsand nonionic surfactants may be used in combination with them.

Useful anionic surfactants include, for example, fatty acid soaps suchas soap stock, sodium laurate and sodium palmitate; higher alkylsulfuric ester salts such as sodium lauryl sulfate and potassium laurylsulfate; alkyl ether sulfuric ester salts such as POE lauryl sulfatetriethanolamine salt and POE lauryl sulfate sodium salt; N-acylsarcosine salts such as sodium lauroyl sarcosine; higher fatty acidamide sulfonate salts such as sodium N-myristoyl N-methyltaurine, sodiumcoconut oil fatty acid methyltauride and sodium lauryl methyltauride;phosphoric ester salts such as sodium POE oleyl ether phosphate and POEstearyl ether phosphate; sulfosuccinic acid salts such as sodiumdi-2-ethylhexyl sulfosuccinate, sodium monolauroyl monoethanolamidepolyoxyethylene sulfosuccinate and sodium lauryl polypropylene glycolsulfosuccinate; alkyl-benzenesulfonic acid salts such as lineardodecylbenzenesulfonic acid sodium salt, linear dodecylbenzenesulfonicacid triethanolamine salt and linear dodecylbenzenesulfonic acid;N-acylglutamic acid salts such as monosodium N-lauroyl glutamate,disodium N-stearoyl glutamate and monosodium N-myristoyl L-glutamate;higher fatty acid ester sulfate salts such as hardened coconut oil fattyacid glycerol sulfate sodium salt; sulfated oils such as Turkey red oil;and POE alkyl ether-carboxylic acids, POE alkyl allyl ether-carboxylicacid salts, α-olefinsulfonic acid salts, higher fatty acid estersulfonicacid salts, secondary alcohol sulfuric ester salts, higher fatty acidalkylolamide sulfuric ester salts, sodium lauroyl monoethanolamidesuccinate, N-palmitoylaspartic acid ditriethanolamine salt and sodiumcaseinate.

Useful cationic surfactants include, for example, alkyltrimethylammoniumsalts (e.g., stearyltrimethylammonium chloride andlauryltrimethylammonium chloride), dialkyldimethylammonium salts (e.g.,distearyldimethylammonium chloride),poly(N,N′-dimethyl-3,5-methylenepiperidinium) chloride, alkylpyridiniumsalts (e.g., cetylpyridinium chloride), alkyl quaternary ammonium salts,alkyldimethylbenzylammonium salts, alkylisoquinolinium salts,dialkylmorpholinium salts, POE alkylamines, alkylamine salts, polyaminefatty acid derivatives, benzalkonium chloride and benzethonium chloride.

Useful ampholytic surfactants include, for example, imidazoline typeampholytic surfactants such as2-undecyl-N,N,N-(hydroxyethylcarboxymethyl)-2-imidazoline sodium saltand 2-cocoyl-2-imidazolinium hydroxide-1-carboxyethyloxy disodium salt;and betaine type surfactants such as2-heptadecyl-N-carboxymethyl-N-hydroxyethylimidazolinium betaine, lauryldimethylaminoacetate betaine, alkyl betaines, amido-betaines andsulfo-betaines.

Useful lipophilic nonionic surfactants include, for example, sorbitanfatty acid esters such as sorbitan mono-oleate, sorbitanmonoisostearate, sorbitan monolaurate, sorbitan monopalmitate, sorbitanmonostearate, sorbitan sesquioleate, sorbitan trioleate, diglycerolsorbitan penta-2-ethylhexylate and diglycerol sorbitantetra-2-ethylhexylate; polyglycerol fatty acid esters such as glycerolmono(cottonseed oil fatty acid) ester, glycerol monoerucate, glycerolsesquioleate, glycerol monostearate, glycerol α,α′-oleate pyroglutamate,and glycerol malate; propylene glycol fatty acid esters such aspropylene glycol monostearate; hardened castor oil derivatives; glycerolalkyl ethers; and polyoxyethylene-methylpolysiloxane copolymers.

Useful hydrophilic nonionic surfactants include, for example, POEsorbitan fatty acid esters such as POE sorbitan mono-oleate, POEsorbitan mono-stearate, POE sorbitan mono-oleate and POE sorbitantetraoleate; POE sorbitol fatty acid esters such as POE sorbitolmonolaurate, POE sorbitol penta-oleate and POE sorbitol monostearate;POE glycerol fatty acid esters such as POE glycerol monostearate, POEglycerol monoisostearate and POE glycerol triisostearate; POE fatty acidesters such as POE mono-oleate, POE distearate, POE dioleate andethylene glycol distearate; POE alkyl ethers such as POE lauryl ether,POE behenyl ether, POE 2-octyldodecyl ether and POE cholestanol ether;POE alkylphenyl ethers such as POE octylphenyl ether, POE nonylphenylether and POE dinonylphenyl; Pluronic type surfactants such as Pluronic;POE-POP alkyl ethers such as POE-POP cetyl ether, POE-POP2-decyltetradecyl ether, POE-POP monobutyl ether, POE-POP hydrogenatedlanolin and POE-POP glycerol ether; tetraPOE-tetra-POP ethylenediaminecondensates such as Tetronic; POE castor oil or POE hardened castor oilderivatives such as POE castor oil, POE hardened castor oil, POEhardened castor oil monoisostearate, POE hardened castor oiltriisostearate, POE hardened castor oil mono-glutamate monoisostearateand POE hardened castor oil maleate; POE beeswax lanolin derivativessuch as POE sorbitol beeswax; alkanolamides such as coconut oil fattyacid diethanolamide, lauric acid monoethanolamide and fatty acidisopropanolamides; and POE propylene glycol fatty acid esters, POEalkylamines, sucrose fatty acid esters, POE nonylphenylformaldehydecondensate, alkyl-ethoxydimethylamine oxides and trioleyl phosphate.

The aforesaid antiseptics include methylparaben, ethylparaben,butylparaben and the like.

The aforesaid sequestering agents include edetic acid sodium salt, EDTAand the like.

The aforesaid water-soluble polymers include natural polymers,semisynthetic polymers, synthetic polymers and inorganic polymers.

Useful natural water-soluble polymers include polymers of vegetableorigin, such as gum arabic, tragacanth gum, galactan, guar gum, carobgum, karaya gum, carrageenan, tamarind gum, xanthane gum, pectin, agar,quince seed (marmelo), algae colloid (brown algae extract), starches(made from rice, corn, potatoes and wheat) and glycyrrhizic acid;polymers of microbial origin, such as xanthane gum, dextran,succinoglucan and pullulan; polymers of animal origin, such as collagen,casein, albumin and gelatin; and the like.

Useful semisynthetic water-soluble polymers include starch-basedpolymers such as dextrin, carboxymethylstarch andmethylhydroxypropylstarch; cellulose-based polymers such asmethylcellulose, nitrocellulose, ethylcellulose,methylhydroxy-propylcellulose, hydroxyethylcellulose, cellulose sulfatedimethyl-dialkyl (12-20)ammonium, hydroxypropylcellulose,carboxymethyl-cellulose sodium (CMC), crystalline cellulose and powderedcellulose; alginic acid-based polymers such as sodium alginate andalginic acid propylene glycol ester; and the like.

Useful synthetic water-soluble polymers include vinyl polymers such aspolyvinyl alcohol, polyvinyl methyl ether, polyvinyl pyrrolidone,carboxyvinyl polymer and alkyl-modified carboxyvinyl polymers;polyoxyethylene polymers such as polyethylene glycols 2000, 4000 and6000; polyoxyethylene-polyoxypropylene copolymers; acrylic polymers suchas sodium polyacrylate, polyethylene acrylate and polyacrylamide;polyethyleneimine; cationic polymers; and the like.

Useful inorganic water-soluble polymers include bentonite, magnesiumaluminum silicate, laponite, hectorite, silicic acid anhydride and thelike.

The aforesaid powder ingredients include inorganic powders such as talc,kaolin, mica, sericite, muscovite, phlogopite, synthetic mica,lepidolite, biotite, lithia mica, vermiculite, magnesium carbonate,calcium carbonate, aluminum silicate, barium silicate, calcium silicate,magnesium silicate, strontium silicate, metallic salts of tungstic acid,magnesium, silica, zeolite, barium sulfate, burned calcium sulfate(calcined gypsum), calcium phosphate, fluorapatite, hydroxyapatite,ceramic powders, metallic soaps (e.g., zinc myristate, calcium palmitateand aluminum stearate) and boron nitride; organic powders such aspowdered polyamide resins (powdered nylons), powdered polyethylene,powdered polymethyl methacrylate, powdered polystyrene, powderedstyrene-acrylic acid copolymer resin, powdered benzoguanamine resin,powdered polytetrafluoroethylene and powdered cellulose; inorganic whitepigments such as titanium dioxide and zinc oxide; inorganic red pigmentssuch as iron oxide (red oxide) and iron titanate; inorganic brownpigments such as β-iron oxide; inorganic yellow pigments such as yellowiron oxide and ochre; inorganic black pigments such as black iron oxide,carbon black and lower titanium oxides; inorganic violet pigments suchas manganese violet and cobalt violet; inorganic green pigments such aschromium oxide, chromium hydroxide and cobalt titanate; inorganic bluepigments such as ultramarine blue and Prussian blue; pearl pigments suchas titanium oxide-coated mica, titanium oxide-coated bismuthoxychloride, titanium oxide-coated talc, colored titanium oxide-coatedmica, bismuth oxychloride and pearl essence; metal powder pigments suchas aluminum powder and copper powder; organic pigments such aszirconium, barium and aluminum lakes including Red No. 201, Red No. 202,Red No. 204, Red No. 205, Red No. 220, Red No. 226, Red No. 228, Red No.405, Orange No. 203, Orange No. 204, Yellow No. 205, Yellow No. 401,Blue No. 404, Red No. 3, Red No. 104, Red No. 106, Red No. 227, Red No.230, Red No. 401, Red No.505, Orange No. 205, Yellow No. 4, Yellow No.5, Yellow No. 202, Yellow No. 203, Green No. 3 and Blue No. 1; naturalpigments such as chlorophyll and β-carotene; and coloring materials suchas Titan Yellow, carthamin and Safflor Red.

The aforesaid ultraviolet protectants include both “ultravioletabsorbers” which are substances capable of absorbing ultraviolet lightchemically, and “ultraviolet screeners” which are substances capable ofscattering and reflecting ultraviolet light by physical action.

Specifically, useful long-wavelength ultraviolet light (UVA) absorbersinclude anthranilic acid type ultraviolet absorbers such as methylanthranilate and homomenthyl-N-acetyl anthranilate; benzophenone typeultraviolet absorbers such as 2,4-dihydroxybenzophenone,2,2-dihydroxy-4-methoxybenzophenone,2,2′-dihydroxy-4,4′-dimethoxybenzophenone,2,2′,4,4′-tetrahydroxy-benzophenone, 2-hydroxy-4-methoxybenzophenone,2-hydroxy-4-methoxy-4′-methylbenzophenone,2-hydroxy-4-methoxybenzophenone-5-sulfonic acid salts,4-phenylbenzophenone, 2-ethylhexyl-4′-phenylbenzophenone-2-carboxylate,2-hydroxy-4-n-octoxybenzophenone and 4-hydroxy-3-carboxybenzophenone;benzotriazole type ultraviolet absorbers such as2,2′-hydroxy-5-methylphenylbenzotriazole,2-(2′-hydroxy-5′-t-octylphenyl)benzotriazole and2-(2′-hydroxy-5′-methyl-phenyl)benzotriazole; dianisoylmethane and4-methoxy-4′-t-butyl-dibenzoylmethane; and the like.

Among these long-wavelength ultraviolet light absorbers,4-methoxy-4′-tert-butyldibenzoylmethane, 2-hydroxy-4-methoxybenzophenoneand 2-hydroxy-4-methoxybenzophenone derivatives (e.g.,2-hydroxy-4-methoxybenzophenone-5-sulfonic acid salts) are preferredlong-wavelength ultraviolet light absorbers having excellent stabilityand effectiveness.

Useful medium-wavelength ultraviolet light (UVB) absorbers includebenzoic acid type ultraviolet absorbers such as p-aminobenzoic acid(hereinafter referred to as PABA), PABA mono-glyceryl ester,N,N-dipropoxyPABA ethyl ester, N,N-diethoxyPABA ethyl ester,N,N-dimethylPABA ethyl ester, N,N-dimethylPABA butyl ester andN,N-dimethylPABA amyl ester; salicylic acid type ultraviolet absorberssuch as dipropylene glycol salicylate, ethylene glycol salicylate,myristyl salicylate, methyl salicylate, amyl salicylate, menthylsalicylate, homomenthyl salicylate, octyl salicylate, phenyl salicylate,benzyl salicylate and p-isopropanolphenyl salicylate; cinnamic acid typeultraviolet absorbers such as octyl cinnamate, ethyl4-isopropylcinnamate, methyl 2,5-diisopropylcinnamate, ethyl2,4-diisopropylcinnamate, methyl 2,4-diisopropylcinnamate, propylp-methoxycinnamate, isopropyl p-methoxycinnamate, isoamylp-methoxycinnamate, octyl p-methoxycinnamate (2-ethylhexylp-methoxycinnamate), 2-ethoxyethyl p-methoxycinnamate, cyclohexylp-methoxycinnamate, ethyl α-cyano-β-phenylcinnamate, 2-ethylhexylα-cyano-β-phenylcinnamate, glycerylmono-2-ethylhexanoyl-p-di-methoxycinnamate, octyl methoxycinnamate,3-methyl-4-[methyl-bis(trimethylsiloxy)silyl]butyl3,4,5-trimethoxycinnamate and mono-ethyl p-dimethoxycinnamate; camphorderivatives such as 3-(4′-methylbenzylidene)-d, 1-camphor,3-benzylidene-d, 1-camphor and5-(3,3-dimethyl-2-norbornylidene)-3-pentyn-2-one; and urocanic acid,ethyl urocanate, 2-phenyl-5-methylbenzoxazole, dibenzalazine and thelike.

Moreover, useful ultraviolet screeners include titanium oxide (TiO₂),talc (MgSiO₂), carmine (FeO₂), bentonite, kaolin, zinc oxide (ZnO) andthe like.

The aforesaid humectants include, for example, polyethylene glycol,propylene glycol, glycerol, 1,3-butylene glycol, hexylene glycol,xylitol, sorbitol, maltitol, chondroitin sulfate, hyaluronic acid,mucoitin sulfate, charonin sulfate acid, atelocollagen, cholesteryl12-hydroxystearate, sodium lactate, bile acid salts,dl-pyrrolidonecarboxylic acid salts, short-chain soluble collagen,diglycerol (EO)PO adduct, Rosa roxburghii extract, yarrow extract andmelilot extract.

The aforesaid active ingredients include whitening agents such asarbutin, vitamin C and its derivatives, kojic acid, placental extract,glutathione and strawberry gerenium extract; anti-inflammatory agentssuch as glycyrrhizic acid derivatives, glycyrrhetic acid derivatives,salicylic acid derivatives, hinokitiol, zinc oxide and allantoin;activators such as royal jelly, photosensitizers, cholesterolderivatives and calf blood extract; blood circulation promoters such asnonylic acid vanillylamide, benzyl nicotinate, β-butoxyethyl nicotinate,capsaicin, gingerone, cantharis tincture, ichthammol, caffeine, tannicacid, α-borneol, tocopheol nicotinate, inositol hexanicotinate,cyclandelate, cinnarizine, tolazoline, acetylcholine, verapamil,cepharanthine and γ-oryzanol; antiseborrheic agents such as sulfur andthianthol; substances effective for various purposes, such asphellodedron bark extract components, coptidis rhizome extractcomponents, lithospermum root extract components, peony root extractcomponents, swertia herb extract components, birch extract components,sage extract components, loquat extract components, ginseng extractcomponents, aloe extract components, mallow extract components, irisextract components, grape extract components, coix seed extractcomponents, sponge gourd extract components, lily extract components,saffron extract components, cnidium rhizome extract components,zingiberis rhizome extract components, syoorengyo extract components,petty white-root extract components, rosemary extract components, garlicextract components, thyme extract components, capsicum extractcomponents, citrus unshiu peel and Japanese angelica root; vitamin Acompounds such as retinol and retinol acetate; vitamin B₂ compounds suchas riboflavin, riboflavin butyrate and flavin adenine dinucleotide;vitamin B₆ compounds such as pyridoxine hydrochloride and pyridoxinedioctanoate; vitamin C compounds such as L-ascorbic acid, L-ascorbicacid dipalmitate, L-ascorbic acid 2-sulfate sodium salt, L-ascorbic acidphosphate and DL-α-tocopherol-L-ascorbic acid diphosphate dipotassiumsalt; pantothenic acid compounds such as calcium pantothenate,D-pantothenyl alcohol, pantothenyl ethyl ether and acetylpentothenylethyl ether; vitamin D compounds such as ergocalciferol andcholecarciferol; nicotinic acid compounds such as nicotinic acid,nicotinic acid amide and benzyl nicotinate; vitamin E compounds such asα-tocopherol, tocopherol acetate, DL-α-tocopherol nicotinate andDL-α-tocopherol succinate; and other vitamins such as vitamin P andbiotin.

As to these active ingredients, they may be extensively compounded intothe compositions of the present invention, so long as the desiredeffects of the present invention are not detracted from thereby.

Since the compositions of the present invention, which have beenprepared in the above-described manner, promote the production oflaminin 5 in human epidermic cells, they can prevent hypofunction of theskin associated especially with structural changes of the basementmembrane, and can promote the potentiation of the skin.

The present invention is more specifically explained with reference tothe following examples. However, these examples are not to be construedto limit the scope of the invention.

Tests for the Ability to Produce Laminin 5

(1) Culture of Epidermic Stratum Corneum Cells

Epidermic stratum corneum cells were isolated from the human preputiumand cultured in an epidermic cell growth medium (KGM) having a lowcalcium concentration. To this culture medium were added bovinepituitary extract and EGF. After the cells were cultured in KGM up tothe fourth generation, and treated with trypsin and EDTA to suspendadherent cells. Then, the culture was filtered to remove cell aggregatesand thereby obtain a homogeneous cell suspension. The cells werecollected by centrifugation and resuspended in DMEM-F12 (2:1)-0.1% BSAso as to give a cell density of 8×10⁴ per ml. 0.5 ml each of this cellsuspension was added to 0.5 ml of the same medium containing a twofoldconcentration of each test sample. Using a 24-well plate, incubation wascarried out at 37° C. for 24 hours. After completion of the incubation,the culture supernatant was transferred to an Eppendorf tube andcentrifuged at 15,000 rpm for 5 minutes. The supernatant was transferredto a new tube and stored at−20° C. till the day for the determination oflaminin 5. In order to solubilize laminin 5 in the cells and bound tothe culture plastics, a tris-HCl buffer (pH 7.4) containing varioussurfactants was added to each well and allowed to stand at −20° C.overnight. On the following day, the mixture was ultrasonicated andfrozen again. On the following day, the mixture was thawed again andcentrifuged at 15,000 rpm for 5 minutes. The supernatant was transferredto a tube and stored at −20° C. till the day for the determination oflaminin 5.

(2) Determination of Laminin 5 by the Sandwich ELISA Method

Laminin 5 present in the culture supernatant and the cell layer wasdetermined by the sandwich ELISA method. A monoclonal antibody (BM165)to the laminin α3 chain of laminin 5 was attached to a solid layer of a96-well ELISA plate. In order to measure laminin 5 in a sandwichedmanner, a monoclonal antibody (6F12) to the laminin β3 chain waspreviously biotinized (b-6F12) and used as the other antibody. In thismethod, only the heterotrimer (α3β3γ2) which can exhibit its functionwas measured, and the heretodimer (β3γ2) was not detected. A sample wasadded to each of the wells in which a 3% gelatin-phosphate buffersolution containing b-6F12 had previously been placed. The final degreeof dilution of the sample in the wells was 1/4 for the culture mediumand 1/10 for the cell layer. After the antigen-antibody reaction wascarried out at 37° C. for 2 hours, the wells were washed, and an avidinHRP (horseradish peroxidase) solution was added thereto and reacted fora period of 30 minutes to 1 hour. After washing, a solution of ABTSserving as a substrate for HRP was added thereto and the absorbance at405 nm was measured with an ELISA plate reader. A calibration curve wasconstructed over a range of 0 to 40 ng/ml.

The yield of laminin 5 was expressed by the sum of the amount liberatedinto the culture medium and the amount remaining in the cell layer.

(3) Results

The results are shown in Tables I and II below.

TABLE I Effect of Lysophosoholipids on the Production of Laminin 5 inEpidermic Stratum Corneum Cells (Comparative tests) Concentration Yieldof laminin 5 (ng) Sample (%) Mean ± Standard deviation Control with noaddition 0 40.3 ± 0.5 Fetal calf serum 1 98.8 ± 4.0 5 135.4 ± 3.5  (Thepresent invention) Relative yield of laminin 5 based Concentration oncontrol with no addition (%) Sample (μg/ml) Mean ± Standard deviationLPC-C12: 0*¹⁾ 5  88.6 ± 5.1 LPC-C14: 0 5 100.8 ± 5.6 LPC-C16: 0 5 104.5± 3.3 LPC-C18: 0 5 112.4 ± 8.0 LPC-C18: 1 5 112.7 ± 6.7 ″ 50 147.4 ± 5.2LPC-C18: 2 5 133.6 ± 1.3 ″ 50 174.0 ± 5.4 LPA-C18: 1 5  225.4 ± 12.5 ″50 254.0 ± 8.7 Lecithin WL-25 5  158.3 ± 14.4 ″ 50  170.0 ± 15.1 KyowaLysolecithin 5 130.6 ± 4.7 ″ 50 140.5 ± 4.7 LIPIDURE 1   144 ± 4.8LIPIDURE 10  163.2 ± 21.0 LIPIDURE 100  240.8 ± 14.2 *¹⁾For comparativepurposes In this table, LPC represents lysophosphatidylcholine. In theexpression “C12: 0” , “12” means the number of carbon atoms contained inR in formula (I), and “0” represents the number of unsaturated doublebonds. The same shall apply to other cases. LPA representslysophosphatidic acid.

5 μg/ml of LPC corresponds to about 10 μM. As to the samples for whichthe yield of laminin 5 is shown only at a concentration 5 μg/ml they mayexhibit an inhibitory effect when used at a high concentration of 50μg/ml.

TABLE II Effect of Soybean-derived Preparations on the Production ofLaminin 5 in Epidermic Stratum Corneum Cells (The present invention)Concentration Yield of laminin 5 (ng) Sample (%) Mean ± Standarddeviation Control with no addition 0 36.7 ± 6.0 Soybean saponin*²⁾0.0001 43.9 ± 3.2 0.001 77.3 ± 3.6 0.01 122.1 ± 3.0  Control with noaddition 0 46.2 ± 6.4 Soybean lecithin*³⁾ 0.0001 51.8 ± 3.2 0.001 69.3 ±2.6 0.01 97.1 ± 2.4 *²⁾ and *³⁾: These samples were obtained from WakoPure Chemical Industries Ltd., Tokyo, as “Saponin, Prepared fromSoybeans” and “Lecithin, Prepared from Soybeans”, respectively.

It can be seen from Tables I and II that the lysophospholipids andsoybean-derived preparations in accordance with the present inventionare effective in promoting the production of laminin 5 to an extentequal to or greater than fetal calf serum which is known to have a cellpotentiating effect.

PREPARATION EXAMPLES Preparation Example 1: Cream

Polyoxyethylene (20 mole addition) cetyl alcohol ether 1.0 Methyl phenylpolysiloxane (20 cs) 2.0 Liquid paraffin 3.02-Hydroxy-4-methoxybenzophenone 5.0 Soybean lysolecithin (ReagentLysolecithin; manu- 0.2 factured by Wako Pure Chemical Industries Ltd.)Propylene glycol 5.0 Glycerol 2.0 Ethyl alcohol 15.0  Carboxyvinylpolymer 0.3 Hydroxypropyl cellulose 0.1 2-Aminomethylpropanol 0.1Antiseptic q.s. Perfume q.s. Ion-exchanged water q.s.

(Preparation Method)

To ion-exchanged water were added propylene glycol, glycerol, ethylalcohol, carboxyvinyl polymer, hydroxypropyl cellulose and2-aminomethylpropanol. This mixture was adjusted to 70° C. by heating(aqueous phase).

Methyl phenyl polysiloxane, liquid paraffin, polyoxyethylene cetylalcohol ether, an antiseptic, 2-hydroxy-4-methoxybenzophenone, soybeanlysolecithin and a perfume were mixed. This mixture was adjusted to 70°C. (oily phase).

The aqueous phase was slowly added to the oily phase so as topre-emulsify this mixture. After the emulsified particles were madeuniform by means of a homomixer, the resulting emulsion was deaeratedand cooled to obtain a cream.

Preparation Example 2: Cream

Polyoxyethylene (20 mole addition) cetyl alcohol ether 1.0 Methyl phenylpolysiloxane (20 cs) 2.0 Liquid paraffin 3.02-Hydroxy-4-methoxybenzophenone 5.0 Lysophosphatidic acid (LPA-C18: 1)0.2 Propylene glycol 5.0 Glycerol 2.0 Ethyl alcohol 15.0  Carboxyvinylpolymer 0.3 Hydroxypropyl cellulose 0.1 2-Aminomethylpropanol 0.1Antiseptic q.s. Perfume q.s. Ion-exchanged water q.s.

(Preparation Method)

To ion-exchanged water were added propylene glycol, glycerol, ethylalcohol, carboxyvinyl polymer, hydroxypropyl cellulose and2-aminomethylpropanol. This mixture was adjusted to 70° C. by heating(aqueous phase).

Methyl phenyl polysiloxane, liquid paraffin, polyoxyethylene cetylalcohol ether, an antiseptic, 2-hydroxy-4-methoxybenzophenone,lysophosphatidic acid and a perfume were mixed. This mixture wasadjusted to 70° C. (oily phase).

Preparation Example 3: Milky lotion

wt. % Cetyl alcohol 1.0 Beeswax 0.5 Vaseline 2.0 Squalane 6.0 Dimethylpolysiloxane 2.0 Ethyl alcohol 5.0 Glycerol 4.0 1,3-Butylene glycol 4.0Soybean lysolecithin (Reagent Lysolecithin; manu- 0.1 factured by WakoPure Chemical Industries Ltd.) Tranexamic acid 1.0 Polyoxyethylene (10)mono-oleate 1.0 Glycerol monostearate 1.0 Quince seed extract (5%aqueous solution) 20.0  Antiseptic q.s. Perfume q.s. Ion-exchanged waterq.s.

(Preparation Method)

Glycerol and 1,3-butylene glycol were added to and mixed withion-exchanged water. This mixture was adjusted to 70° C. by heating(aqueous phase). Cetyl alcohol, beeswax, vaseline, squalane, dimethylpolysiloxane, soybean lysolecithin, tranexamic acid, polyoxyethylene(10) mono-oleate, glycerol monostearate and an antiseptic were mixed.This mixture was adjusted to 70° C. by heating (oily phase). The aqueousphase was added to the oily phase so as to pre-emulsify this mixture.Then, quince seed extract and ethyl alcohol were added to the resultingemulsion, followed by stirring. After the emulsified particles were madeuniform by means of a homomixer, the resulting emulsion was deaerated,filtered, and cooled to obtain a milky solution.

Preparation Example 4: Milky lotion

wt. % Cetyl alcohol 1.0 Beeswax 0.5 Vaseline 2.0 Squalane 6.0 Dimethylpolysiloxane 2.0 Ethyl alcohol 5.0 Glycerol 4.0 1,3-Butylene glycol 4.0Soybean lysolecithin (LIPIDURE NC-22; manu- 0.1 factured by Nippon Oil &Fats Co., Ltd.) Tranexamic acid 1.0 Polyoxyethylene (10) mono-oleate 1.0Glycerol monostearate 1.0 Quince seed extract (5% aqueous solution)20.0  Antiseptic q.s. Perfume q.s. Ion-exchanged water q.s.

(Preparation Method)

Glycerol and 1,3-butylene glycol were added to and mixed withion-exchanged water. This mixture was adjusted to 70° C. by heating(aqueous phase). Cetyl alcohol, beeswax, vaseline, squalane, dimethylpolysiloxane, soybean lysolecithin, tranexamic acid, polyoxyethylene(10) mono-oleate, glycerol monostearate and an antiseptic were mixed.This mixture was adjusted to 70° C. by heating (oily phase). The aqueousphase was added to the oily phase so as to pre-emulsify this mixture.Then, quince seed extract and ethyl alcohol were added to the resultingemulsion, followed by stirring. After the emulsified particles were madeuniform by means of a homomixer, the resulting emulsion was deaerated,filtered, and cooled to obtain a milky solution.

The aqueous phase was slowly added to the oily phase so as topre-emulsify this mixture. After the emulsified particles were madeuniform by means of a homomixer, the resulting emulsion was deaeratedand cooled to obtain a cream.

Preparation Example 5: Cream

Polyoxyethylene (20 mole addition) cetyl alcohol ether 1.0 Methyl phenylpolysiloxane (20 cs) 2.0 Liquid paraffin 3.02-Hydroxy-4-methoxybenzophenone 5.0 Soybean saponin (Saponin, Preparedfrom Soybeans; 0.2 manufactured by Wako Pure Chemical Industries Ltd.)Propylene glycol 5.0 Glycerol 2.0 Ethyl alcohol 15.0  Carboxyvinylpolymer 0.3 Hydroxypropyl cellulose 0.1 2-Aminomethylpropanol 0.1Antiseptic q.s. Perfume q.s. Ion-exchanged water q.s.

(Preparation Method)

To ion-exchanged water were added propylene glycol, glycerol, ethylalcohol, carboxyvinyl polymer, hydroxypropyl cellulose and2-aminomethylpropanol. This mixture was adjusted to 70° C. by heating(aqueous phase).

Methyl phenyl polysiloxane, liquid paraffin, polyoxyethylene cetylalcohol ether, an antiseptic, 2-hydroxy-4-methoxybenzophenone, soybeansaponin and a perfume were mixed. This mixture was adjusted to 70° C.(oily phase).

The aqueous phase was slowly added to the oily phase so as topre-emulsify this mixture. After the emulsified particles were madeuniform by means of a homomixer, the resulting emulsion was deaeratedand cooled to obtain a cream.

Preparation Example 6: Cream

Polyoxyethylene (20 mole addition) cetyl alcohol ether 1.0 Methyl phenylpolysiloxane (20 cs) 2.0 Liquid paraffin 3.02-Hydroxy-4-methoxybenzophenone 5.0 Soybean lecithin (Lecithin, Preparedfrom Soybeans; 0.2 manufactured by Wako Pure Chemical Industries Ltd.)Propylene glycol 5.0 Glycerol 2.0 Ethyl alcohol 15.0  Carboxyvinylpolymer 0.3 Hydroxypropyl cellulose 0.1 2-Aminomethylpropanol 0.1Antiseptic q.s. Perfume q.s. Ion-exchanged water q.s.

(Preparation Method)

To ion-exchanged water were added propylene glycol, glycerol, ethylalcohol, carboxyvinyl polymer, hydroxypropyl cellulose and2-aminomethylpropanol. This mixture was adjusted to 70° C. by heating(aqueous phase).

Methyl phenyl polysiloxane, liquid paraffin, polyoxyethylene cetylalcohol ether, an antiseptic, 2-hydroxy-4-methoxybenzophenone,lysophosphatidic acid and a perfume were mixed. This mixture wasadjusted to 70° C. (oily phase).

The aqueous phase was slowly added to the oily phase so as topre-emulsify this mixture. After the emulsified particles were madeuniform by means of a homomixer, the resulting emulsion was deaeratedand cooled to obtain a cream.

INDUSTRIAL APPLICABILITY

Since the lysophospholipids and soybean-derived preparations inaccordance with the present invention can promote the production oflaminin 5 in epidermic cells, compositions containing them as activeingredients and their use can be utilized especially in the fields ofcosmetics and dermatology.

What is claimed is:
 1. A method for potentiating epidermic cells of amammal which comprises administering a compound of the following formula(I) or a mixture of two or more of such compounds to the mammal in anamount sufficient to promote the production of laminin 5 in epidermiccells:

wherein R is a C₁₉₋₂₂ fatty acid residue optionally having up to sixunsaturated double bonds, A is a hydrogen atom or a —CH₂CH₂N⁺(CH₃)₃group, and M is a hydrogen atom or an alkali metal atom, but is ahydrogen atom when A is a —CH₂CH₂N⁺(CH₃)₃ group.
 2. A method forpotentiating epidermic cells of a mammal which comprises administering apreparation derived from soybeans to the mammal in an amount sufficientto promote the production of laminin 5 in epidermic cells, wherein thepreparation derived from soybeans is an organic solvent extract fractioncontaining both a soybean saponin fraction and a soybean lecithinfraction derived from soybeans and having an activity for promoting theproduction of laminin 5 in human keratinized epidermic cells.
 3. Amethod for potentiating epidermic cells of a mammal which comprisesadministering a preparation derived from soybeans to the mammal in anamount sufficient to promote the production of laminin 5 in epidermiccells, wherein the preparation derived from soybeans is an organicsolvent extract fraction containing a soybean lecithin fraction derivedfrom soybeans and having an activity for promoting the production oflaminin 5 in human keratinized epidermic cells, with the proviso thatthe preparation does not contain a soybean saponin fraction.